The goal of this project is to develop an anti-Pseudomonas aeruginosa vaccine using a novel adenovirus (Ad)-based gene transfer vector that: codes for the P. aeruginosa outer membrane protein F (OprF);has been enhanced to incorporate an RGD sequence in the fiber to enhance delivery to dendritic cells in vivo via ?v?3.5 integrins;and incorporates Epi8, a conserved, highly immunogenic OprF epitope, into the hexon outer loop. Aim 1 focuses on using this strategy with an Ad5-based vaccine (Ad5cuOprF.RGD.Epi8). We expect that it will elicit high levels of anti-P. aeruginosa antibodies in blood and lung, and that the Epi8 epitope on the capsid will allow boosting, even in the presence of immunity against the Ad induced by the 1st immunization. After completing the toxicology testing, and gaining regulatory approval, we will carry out a clinical trial with Ad5cuOprF.RGD.Epi8 in normal adult volunteers. In aim 2, we will carry out studies with AdC7cuOprF.RGD.Epi8, a 2nd generation anti-P. aeruginosa vaccine that is similar to Ad5CuOprF.RGD.Epi8, but based on the non-human primate AdC7 serotype, a serotype that should be effective even in the presence of pre-existing anti-Ad5 immunity. The specific aims of the project are: Aim 1 - To test the hypothesis that administration of Ad5cuOprF.RGD.Epi8, a human Ad serotype 5 adenovirus that codes for the P. aeruginosa antigen OprF and is further modified with a dendritic cell targeting motif RGD in the fiber and the OprF Epi8 epitope in the hexon, is safe and will evoke robust anti-Pseudomonas opsonizing antibodies in blood and lung epithelial lining fluid of healthy individuals;Aim 2 - To examine the hypothesis that effective anti-P. aeruginosa immunity can be achieved in the context of anti-Ad5 immunity with AdC7cuOprF.RGD.Epi8, a vector identical to Ad5cuOprF.RGD.Epi8 but based on AdC7, a non-human primate adenovirus against which humans do not have immunity. Infections of the respiratory tract with Pseudomonas aeruginosa is the major cause of morbidity and mortality in individuals with cystic fibrosis. A marketed vaccine for prevention of infection with Pseudomonas aeruginosa is not available, therefore a successful vaccine strategy would have profound effects on individuals with CF.